PROJECT SUMMARY (Description) Stroke is a common and devastating complication of Sickle Cell Anemia (SCA), and the current management relies on transfusion therapy without the use of thrombolytic agents. Humanized sickle mice (knock-in/out mice that express the human ?, ?, and sickle-? hemoglobin genes) are powerful tools to develop treatments of SCA complications, but their use in stroke research has been limited by the fragility of these mutant mice. To overcome these challenges, we have developed a modified photothrombosis model that is well suited for the research of acute stroke therapy in humanized sickle mice. In this project, we will assess the risks and benefits of two promising therapeutics?tissue plasminogen activator (tPA)-mediated thrombolysis and Gardos/KCa3.1 channel blockers?and test for their additive or synergistic benefits in acute sickle cell stroke therapy. Positive outcomes of this project will provide insights into the clinical development of lytic stroke therapy in SCA. Aim 1: Determine the safety doses and therapeutic window of tPA-thrombolysis in sickle mice. We will determine the safety dose-range and therapeutic window of tPA treatment for the thrombin- photothrombosis stroke model between sickle (SS) and non-sickle (AA) mice. Aim 2: Determine the risk and benefits of combined tPA/KCa3.1 blocker therapy in sickle mice. With the key role of calcium-dependent potassium channel (Gardos/KCa3.1) in RBC dehydration/sickling, and microglia/macrophage immune response, we will compare the risks and benefits of tPA, TRAM-34 (a potent KCa3.1 blocker), and tPA/TRAM-34 treatment of thrombin/photothrombosis stroke in SS mice.